![Loss of p53 in human HCT116 colon carcinoma cells alters the behavior of multiple cellular pathways.(A) Cell viability analysis by AnnexinV/PI staining on TP53+/+ and TP53−/− HCT116 cells in 5% O2 after 0.2 µg/ml doxorubicin or 0.1% FBS treatment for 48 h. n = 3 independent experiments for each genotype. Data are presented as mean ± SD; *, P < 0.05, two-way ANOVA, Sidak’s multiple comparison posttest. (B) Proliferation analysis of TP53+/+ and TP53−/− HCT116 cells measured by cell counting starting 24 h after plating. n = 3 independent experiments. Data are presented as mean fold change in cell number ± SD. (C) Ploidy analysis in TP53+/+ and TP53−/− HCT116 cells showing percentage of cells with >4N DNA content by PI staining. n = 3 independent experiments per genotype; data represent mean ± SD; *, P < 0.05, unpaired t test. (D and E) [U13C]glucose tracing on TP53+/+ and TP53−/− HCT116 cells showing glycolytic (D) and TCA cycle (E) intermediates. Data represent mean ± SD of n = 3 cell lines per sgRNA; *, P < 0.05; **, P > 0.005; ***, P < 0.001; ****, P < 0.0001, one-way ANOVA, Tukey’s multiple comparison test. (F) Citrate:lactate mass heavy ion ratio of TP53+/+ and TP53−/− HCT116 cells. Data represent mean ± SD of n = 2 samples per cell line; Student’s t test. (G) Representative images of TP53+/+ and TP53−/− HCT116 cells grown in a 3D collagen matrix. WGA stains cell membranes, and DAPI marks nuclei. Scale bar, 0.2 mm. (H) Percentage of invading colonies in 3D collagen assay. Data are mean ± SD, n = 2 independent experiments per genotype, P = 0.073, unpaired t test. All HCT116 experiments were performed in 5% O2.](https://cdn.rupress.org/rup/content_public/journal/jcb/219/11/10.1083_jcb.201908212/4/jcb_201908212_figs3.png?Expires=1771142762&Signature=5OtZp2HAN2k19A4DWgT-MYFsHkemwnsGWq1ONguvRUzLHHx6uoLu4F2FhfdICxqAt8XKqyct59-ua6ijdYEqfN1M5kbDFj1U6skd7tVTULX7rMUCOOC2XfWJWS3zzFY9htob7ain08cmHqiPDYDIwwQ331ueweBTIYl9jRJnP2TlCNy3aBLhcVTPDv-wtuiqBCY0t5d875phjPwlgcyrd8yiU6o1BOTWByYfphnysyuomPXt-hDaZXW9d9CCtmJL1tlmjmKSt353JQKWtt4WyILM~nc7lt7YTOF2l~qgBDgi4SQzrYJUNS9543cTsAcZbRyHzS~1kXOSQcwpYC~l6A__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Loss of p53 in human HCT116 colon carcinoma cells alters the behavior of multiple cellular pathways. (A) Cell viability analysis by AnnexinV/PI staining on TP53+/+ and TP53−/− HCT116 cells in 5% O2 after 0.2 µg/ml doxorubicin or 0.1% FBS treatment for 48 h. n = 3 independent experiments for each genotype. Data are presented as mean ± SD; *, P < 0.05, two-way ANOVA, Sidak’s multiple comparison posttest. (B) Proliferation analysis of TP53+/+ and TP53−/− HCT116 cells measured by cell counting starting 24 h after plating. n = 3 independent experiments. Data are presented as mean fold change in cell number ± SD. (C) Ploidy analysis in TP53+/+ and TP53−/− HCT116 cells showing percentage of cells with >4N DNA content by PI staining. n = 3 independent experiments per genotype; data represent mean ± SD; *, P < 0.05, unpaired t test. (D and E) [U13C]glucose tracing on TP53+/+ and TP53−/− HCT116 cells showing glycolytic (D) and TCA cycle (E) intermediates. Data represent mean ± SD of n = 3 cell lines per sgRNA; *, P < 0.05; **, P > 0.005; ***, P < 0.001; ****, P < 0.0001, one-way ANOVA, Tukey’s multiple comparison test. (F) Citrate:lactate mass heavy ion ratio of TP53+/+ and TP53−/− HCT116 cells. Data represent mean ± SD of n = 2 samples per cell line; Student’s t test. (G) Representative images of TP53+/+ and TP53−/− HCT116 cells grown in a 3D collagen matrix. WGA stains cell membranes, and DAPI marks nuclei. Scale bar, 0.2 mm. (H) Percentage of invading colonies in 3D collagen assay. Data are mean ± SD, n = 2 independent experiments per genotype, P = 0.073, unpaired t test. All HCT116 experiments were performed in 5% O2.
